PROJECT SUMMARY Fibrinogen is essential for normal blood coagulation and is an integral component of inflammatory pathways. These two processes are deeply intertwined in the development of thrombotic and atherosclerotic diseases. We can better understand the biological and pathophysiological actions of fibrinogen by functionally characterizing the genomic contribution to circulating fibrinogen levels. In the Cohorts for Heart and Aging Research in Genomic Epidemiology (CHARGE) consortium, we carried out genome-wide association studies (GWAS) in search of genetic determinants of fibrinogen levels. These studies involved tens of thousands of individuals and identified variants within fibrinogen structural genes, as well as 42 other significantly associated loci. However, association studies do not explain which genes at these loci functionally influence fibrinogen levels. It is also important to determine whether genes at these loci act through inflammatory pathways. The goal of this project is to leverage our expertise in genomic studies and functional biology to generate new biological knowledge about the genomic regulation of fibrinogen and to characterize the relationship between fibrinogen and thrombotic and atherosclerotic disease. Our study design enables information exchange between functional biologists with expertise in coagulation biochemistry and thrombosis pathophysiology, and genetic epidemiologists from the CHARGE consortium. The interdisciplinary team will carry out three specific aims. First, we will use siRNA gene silencing to interrogate genes at 42 loci identified by GWAS to be associated with fibrinogen levels, determine their effect on fibrinogen transcription, translation, and secretion, and establish whether these genes modulate fibrinogen levels via an inflammatory (IL6-STAT3) pathway. Second, we will perform an epigenome-wide association study to examine the association between fibrinogen levels and blood methylation levels at CpG sites across the genome. These results are integrated with genetic data to identify genetic variants associated with CpG sites (meQTLs) that are also associated with fibrinogen levels. Candidate genes at fibrinogen-associated meQTLs will also be evaluated using the proposed functional biological methods. Third, we will assess the causal relationship between fibrinogen and thrombotic and atherosclerotic disease using a Mendelian Randomization approach. Knowledge gained from the functional biology experiments will be used to refine the genetic instrument for fibrinogen and additionally create an instrument that is composed only of variants related to the IL6-STAT3 pathway. This team science approach is innovative in its experimental design and in its potential to reveal important, new information impacting human health by translating results of genomic association studies into a clear understanding of fibrinogen's role in thrombotic and atherosclerotic disease.